Abstract

Arginine-rich cell-penetrating peptides (AR-CPPs) are very promising tools for the delivery of therapeutic macromolecules such as peptides, proteins, and nucleic acids. These peptides allow efficient internalization of the linked cargos intracellularly through the endocytic pathway. However, when linked to bulky cargos, entrapment in the endocytic vesicles is a major limitation to the application of these peptides in cytosolic delivery. Attachment of a compatible endosomal escape device is, therefore, necessary to allow cytosolic delivery of the peptide-attached cargo. This review presents different endosomal escape devices currently in application in combination with AR-CPPs. Applications of fusogenic lipids, membrane-disruptive peptides, membrane-disruptive polymers, lysosomotropic agents, and photochemical internalization to enhance the cytosolic delivery of AR-CPPs-attached cargos are presented. The properties of each system and its mechanism of action for the enhancement of endosomal escape are discussed, together with its applications for the delivery of different macromolecules in vitro and, if applicable, in vivo.

PEGylation of liposomes linked to AR-CPPs hinders fusion between the liposomes and endosomal membrane. Despite the electrostatic attraction between the AR-CPPs on the liposome surface and the negatively charged components in endosomal membrane, fusion does not take place due to steric hindrance by the bulky hydrophilic PEG chains

Model for the role of AR-CPPs in the fusion between liposome and endosomal membrane. Steps of the fusion between liposome modified with octaarginine, a model AR-CPP, and endosome are summarized. a Recognition of the site of fusion between the liposome and endosomal membrane through electrostatic attraction between the cationic AR-CPP (R8) and the anionic components in the endosomal membrane. b Close apposition of the membranes, aided by the electrostatic attraction and neutralization of the positive charges on R8 by the endosomal membrane anionic components. c Fusion or coalescence of the endosomal membrane and the liposome. Fusion is guided by insertion of the neutralized R8 into the endosomal membrane and movement inwards under the influence of difference in endosomal transmembrane potential. d Fusion pore is formed following lipid mixing between the two membranes and resulting in the release of the liposome cargo into the cytosol

Endosomal escape of PEI polyplexes by proton sponge effect. After uptake, PEI polyplexes in the endosome act as proton sponge through protonation of the secondary and tertiary amine groups in the PEI polymer chain. Accumulation of protons together with their counter ions in the endosome stimulates entrance of water from the cytosol to balance the high osmotic pressure inside the endosome. Swelling of the endosome in the presence of PEI eventually leads to endosome bursting and release of PEI polyplexes to the cytosol